The field of reproductive endocrinology has struggled with many aspects of the regulation of the gonadotropin beta subunit genes, in large part due to the lack of a well-differentiated beta-subunit expressing cell line. The development of the LbetaT2 cell line which was demonstrated to express and secrete LH (1) has now been shown to also express and secrete FSH(2) and a second cell line, the FSHtsT5 line, secretes FSH as well. These two cell lines are the first to express all three gonadotropin genes and preliminary studies indicate that they exhibit differential regulation of those genes. However, the applicability of these two cells lines to study of the gonadotropin genes is dependent upon how well they recapitulate normal gonadotroph physiology and that has not yet been established. We hypothesize that cells of the LbetaT2 and FSHtsT5 lines retain elements of differentiated gonadotroph function and propose to evaluate them as models of mature, differentiated gonadotrophs. We plan on measuring their expression of gonadotroph-specific genes important for the expression of the gonadotropin subunits including activin, inhibin, follistatin and their receptors. This will not only confirm their appropriateness for study of LH and FSH, but also the study of the intracellular signalling of activin and the complex interactions between these gonadal peptides. We also propose to evaluate the response of these cell lines to known activators of gonadotroph function including GnRH, activin, inhibin, and the gonadal steroids. This is essential groundwork for more complex studies of factors which contribute to the differential regulation and secretion of LHbeta, FSHbeta and alpha-subunit. In addition, this project supports the Principal Investigator's Career Development Plan in conjuction with K08-DK 02477 with technical support for continued scientific productivity and further specialized research technique training.